The Cas9-guide RNA complex (sphere) slides along the DNA helix searching for a PAM site whose sequence matches the guide RNA. A match triggers editing: classic Cas9 makes a double-strand cut, base editors chemically convert one letter without cutting, and prime editors rewrite a short sequence directly.
P(on-target edit) = specificity
P(off-target edit) = (1 - specificity) × off_target_site_density
- Editing mode — Cas9 Cut shows a double-strand break; Base Edit shows a single-base color swap with no break; Prime Edit shows a localized rewritten segment.
- Guide RNA specificity — probability the complex correctly distinguishes the intended target from similar decoy sequences.
- Scan speed — how fast the Cas9 complex moves along the DNA searching for matches.
Real-world application: this on-target vs. off-target trade-off is precisely what determines whether an approved CRISPR therapy (e.g. for sickle cell disease) is judged safe and effective in clinical trials.